Our long-term goal is to understand the cancer preventive activities of tea constituents and their applications to human cancer prevention. The cancer preventive activity of tea polyphenols has been demonstrated in animal models, but the mechanisms of action are not clearly understood. This project aims to elucidate the mechanisms of lung cancer prevention by (-)-epigallocatechin-3-gallate (EGCG), the major green tea polyphenol. Our goal is to establish an integrated in vitro and in vivo experimental system to test our central hypothesis that EGCG inhibits lung carcinogenesis by binding to specific target molecules which are vital to carcinogenesis and this binding triggers alterations in signal transduction pathways that lead to growth inhibition and apoptosis of premalignant and malignant cells. In order to effectively test our hypothesis, we plan to use both in vitro and in vivo experimental systems. The specific aims are as follows: 1. Establish and compare in vivo (xenograft & allograft) and in vitro experimental systems with lung cancer cell lines and study the mechanisms of action of EGCG. Lung cancer cell lines H1299 (human) and CL13 and CL30 (mouse) will be studied in culture and in xenografts/allografts. The effects of EGCG treatment on tumor growth, apoptosis, angiogenesis, and related molecular changes (e.g., ERK1/2, AKT, ASK1, JNK, VEGF, and oxidative stress parameters) will be compared in vivo vs. in vitro. 2. Identify direct molecular targets of EGCG by affinity-proteomics and functional studies. To identify direct molecular targets for EGCG, we will use affinity chromatography and mass spectrometry in collaboration with Dr. Zigang Dong at the University of Minnesota. The functional roles of the putative EGCG target proteins in mediating the action of EGCG will be studied with loss or gain of function experiments in engineered cells and in a xenograft/allograft model established in Aim 1. 3. Elucidate the mechanisms of cancer prevention by EGCG in a 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung carcinogenesis model in A/J mice. Adenoma-bearing mice will receive long- or short-term treatment with EGCG. The cellular and molecular changes related to the inhibition of tumor progression will be investigated using immunohistochemistry, Western blots, and other approaches. We will test specific hypotheses developed based on results from Aims 1 and 2 concerning putative EGCG molecular targets and related molecular events. A better understanding of the mechanisms of the cancer preventive action of EGCG and the dose-response relationship will help us to design future lung cancer prevention trials in humans, for example, in ex-smokers. Project Narrative: Green tea has been shown to have cancer preventive activity in animal models. This project aims to study the lung cancer preventive mechanisms of the major green tea polyphenol, (-)-epigallocatechin-3-gallate (EGCG). The results will help us to design future lung cancer prevention trials in humans and to assess the effects of tea consumption on human lung cancer.